Psychostimulant-induced dendritic spine plasticity in brain reward circuits might be a cellular mechanism of learning and enduring memory associated with addictive behaviors. We hypothesize that two types of dendritic spine formation (silent spines and mature spines) may occur during chronic exposure, withdrawal and re-exposure to psychostimulants. Cell type- and region-specific studies of dendritic spines and of the role of Cdk5 and WAVE1 in the molecular mechanisms underlying the two steps of spine formation are designed with novel and innovative approaches to achieve the following aims. In Aim I, cell type- and region-specific analysis of 1) dendritic spine morphology of medium-sized spiny neurons (MSNs) and of 2) the levels of glutamate receptor expression in dendritic spines will characterize the structure and function of dendritic spines. We will use Bac-transgenic mice expressing a cell type-specific fluorescent marker protein to identify specific MSNs. We will generate three-dimensional images of dendritic spines from two types of MSNs in subregions of striatum (core and shell of nucleus accumbens and dorsal striatum) during chronic exposure, withdrawal and re-exposure to cocaine. Various morphological parameters of dendritic spines will be analyzed with automatic software. In Aim II and III, we will use D1- or D2-MSNs-specific Cdk5 or WAVE1 knockout mice. We will also restore the expression of Cdk5 or WAVE1 only in the specific types of MSNs in specific regions of striatum (nucleus accumbens or dorsal striatum) by injection of conditional AAV-floxed-STOP signal-floxed-Cdk5 or WAVE1 (wild-type or phosphorylation site mutants). With these knockout and add-back Cdk5 or WAVE1 mice, we will study the roles of Cdk5, WAVE1 and phosphorylation of WAVE1 in (a) spine morphology, (b) glutamate receptor trafficking, (c) electrophysiology and (d) behavior. In Aim III.C, we will characterize WAVE1 phosphorylation at tyrosine sites by TrkB, a receptor of mature BDNF, in the actions of psychostimulants. In addition, we will carry out collaborative studies with Project 2 in the analysis of dendritic spines in genetically modified animal models (e.g. CK1 overexpressing mice). We will also collaborate with Project 3 in studies of the role of PP2A in WAVE1 regulation and of the role of Rap1GAP in dendritic spine morphogenesis.